Nucleic acids, either DNA or RNA, single-stranded or double-stranded, are the most fundamental and important class of biomolecules in a living cell. DNA encodes the genetic information that passes from generation to generation. Through transcription, the coded information is transferred to mRNA, which binds to ribosome (specific ribosomal RNA and protein complex). With the assistance of tRNA, which contains an anticodon and a specific amino acid, the carried information of mRNA is translated into a precise sequence of a polypeptide of 20 amino acids. Folding of the polypeptide into a well-defined three-dimensional structure gives the protein. Many classes of protein act as building blocks, enzymes, and regulation factors. Together with other classes of biomolecules, proteins are responsible for the buildup and proper function of a living cell.
Since nucleic acids carry multiple negatively charged phosphate functional groups, they are polyanions. Under physiological conditions, poly(aspartic acid) and poly(glutamic acid) form polycarboxylates, which are also polyanions. On the other hand, polylysine, polyarginine, and polyhistidine (in an acidic aqueous solution) carry multiple positive charges, and are considered polycations. Many proteins, when the solution pH is not at their isoelectric point (pI) value, carry net positive or negative multiple charges. In light of the above, methods that can detect and characterize biomolecules with multiple charges are of great importance, which can not only help us to understand how the cell functions, assist biological/biochemical research, but may also provide ways to facilitate biomedical research, clinical diagnosis, and new drug development.
The intriguing structural and bonding properties of square-planar d8 or d10 metal complexes have attracted long-standing interest, and more so recently with the growing interest in the spectroscopic properties associated with this class of metal complexes. These metal complexes are known to display a strong tendency towards the formation of highly-ordered extended linear chains or oligomeric structures in the solid state. The extent of the metal-metal interaction and the π . . . π stacking of the aromatic ligand have led to the observation of interesting spectroscopic and luminescence properties, and recent reports based on the utilization of these observations for molecular recognition, chemosensing, and optoelectronic applications have been made. (17, 23, 27)
A representative example of the class of the aforementioned d8 or d10 metal complexes is the alkynylplatinum(II) terpyridyl complexes (25, 26, 28). By changing the solvent polarity, or using a polyelectrolyte (a polyanion), namely polyacrylate, the d8 or d10 metal complexes are induced to aggregate and self-assemble, thereby creating observable dramatic changes in the UV/vis and emission spectra.
There are a number of assay methods available nowadays for the detection and characterization of multiple-charged biomolecules. However, most of the commonly used existing methods require sophisticated analytical techniques and expensive instrumentations. Many of these methods require labeling with a detectable group, which can be a radioisotope or a fluorescent substance, as well as hybridization procedures for nucleic acid detection. Hence, such methods usually demand high financial cost and are technically complicated and time-consuming.
The present invention provides a novel label-free assay method to sense and characterize multiple-charged biomolecules. Binding of the charged d8 or d10 metal complex to the biomolecule carrying opposite charges induces aggregation and self-assembly of the metal complex, and hence gives rise to remarkable UV/vis, emission, and CD intensity changes. The assay not only provides a means to detect the presence of multiple-charged biomolecules, but can also be used to study their secondary structure and structure/conformation changes.